Perennial endemic and potentially pandemic influenza virus infections constitute a major global health problem. It is known that the adaptive CD8 T cell response is vital for clearance of acute influenza virus infection, and that antibodies are important for protection against recurrent infections. Relatively little is understood about how the innate phase of immunity impacts the success of these adaptive components. Natural killer (NK) cells are cytotoxic lymphocytes that respond in the earliest phase of viral infections to control viral spread by direct killing of infected cells, and by secretion of potent cytokines (e.g., interferon-gamma, IFN-g) that enhance the priming of a Th1 adaptive immune response. We have found that depletion of NK cells prior to influenza A virus (IAV) infection results in exacerbated disease, and that these mice may be rescued from early demise by reconstitution of NK cell immunity using adoptively transferred lung NK cells. The long-range goal for this project is to elucidate the molecular mechanisms through which NK cells mediate this protection. The experimental design is one that provides new information about the integrated immune response operating during the early innate and subsequent transitional phases of acute IAV infection-from the recruitment of NK cells into the lungs and draining lymph nodes, through the effector phase of NK cell activation, and ultimately to the establishment of a pulmonary CD8 T cell response. We will exploit novel in vivo and in vitro experimental systems including our NK cell depletion-reconstitution in BALB.B6-CT6 (NK1.1+) mice and ex vivo intact lymph node culture to isolate NK cell responses and functions through three Specific Aims: 1) a determination of the mechanisms of: a) homeostatic NK cell homing to the lungs following adoptive transfer, and b) pulmonary NK cell accumulation during acute IAV infection (proliferation, differential apoptosis, and/or recruitment);2) a determination of the mechanism of NK cell-mediated protection against lethal disease during IAV infection-role of interferon-g, perforin, Fas ligand and TRAIL in control of peak viral load and the virus-specific CD8 T cell response;and 3) a determination of the contributions from proliferation, differential apoptosis, and CXCR3- dependent recruitment to NK cell accumulation in the draining lymph node during acute IAV infection and potential regulation by migratory pulmonary DC and/or macrophages. The results from this project will expand our understanding of the integrated immune response to a major respiratory pathogen, and will impact the design of vaccines in order to exploit the beneficial contributions of NK cells. PUBLIC HEALTH RELEVANCE: Influenza virus infections exact an enormous toll on human populations: seasonal endemic infections result in >35,000 deaths annually in the US, while recurrent pandemic infections constitute a major global threat. The results from this project will enhance our understanding of how innate immunity works to control spread of the acute infection and initiate clearance by the adaptive immune system, and how dysregulation in the innate phase may occur (and be effectively treated) in severe influenza infections. Finally, the findings will impact the design of new vaccination strategies in order to exploit the contributions from natural killer cells.